LID scientists are collaborating with scientists from MedImmune under a CRADA to generate candidate vaccines against avian influenza viruses of each subtype. The vaccines were generated using plasmid based reverse genetics and each contains the hemagglutinin and neuraminidase genes from an avian influenza virus and six internal gene segments from the AA ca virus. Since 1968, human H3N2 influenza viruses have been circulating worldwide, and the population has been infected by and/or vaccinated against these viruses. However, H3 subtype influenza viruses have been isolated from humans, pigs, horses, dogs, cats, seals, and numerous avian species, and these viruses are antigenically and genetically distinct from human H3 viruses. Swine, equine, and avian H3 influenza viruses all have crossed the species barrier to mammals, including humans, dogs, and seals. Very little information is available on the kinetics of replication and cross-reactivity of the immune response of swine, equine, and avian H3 influenza viruses in experimental animal models. Based on the emergence of a swine-origin H1N1 virus as a pandemic strain in 2009 despite the ongoing circulation of human H1viruses, we believe it is necessary to consider the development of H3 animal influenza vaccines. We selected 11 swine, equine, and avian H3 influenza viruses and evaluated their kinetics of replication and ability to induce a broadly cross-reactive antibody response in mice and ferrets. The swine and equine viruses replicated well in the upper respiratory tract of mice. With one exception of an avian virus that replicated poorly in the lower respiratory tract, all of the viruses replicated in mouse lungs. In ferrets, all of the viruses replicated well in the upper respiratory tract, but the equine viruses replicated poorly in the lungs. Extrapulmonary spread was not observed in either mice or ferrets. No single virus elicited antibodies that cross-reacted with viruses from all three animal sources. Avian and equine H3 viruses elicited broadly cross-reactive antibodies against heterologous viruses isolated from the same or other species, but the swine viruses did not. We selected an equine and an avian H3 influenza virus for further development as vaccines.